Background Rheumatoid arthritis synovial fibroblasts (RASF) are characterized by reduced DNA methylation which is supposed to play a role in development of their stably activated destructive phenotype. Agents restoring DNA methylation have been proposed as a promising therapeutic strategy for the treatment of RA. One of the potential agents to reverse hypomethylation is methyl donor S-Adenosyl-L-Methionine (SAMe). A soy phytoestrogen genistein has been shown to act similar to methyl donor supplementation on certain cells lines. IL18, an inflammatory cytokine, produced by RASF and playing an important role in RA inflammation and angiogenesis, has recently been considered as an attractive target for RA treatment. To date, there have been no studies investigating the effects of SAMe and genistein on global DNA methylation and IL-18 production by RASF.
Objectives To evaluate in vitro effects of genistein and SAMe on global DNA methylation and IL-18 production by activated RASF.
Methods RASF were isolated from synovial tissue obtained during joint replacement surgery from 3 RA patients. The cells were stimulated with 10 ng/ml IL-1β and then SAMe (25, 50, 100 ug/ml) or genistein (5, 10, 20 ug/ml) were added to cultures. Global DNA methylation was measured by flow cytometry using monoclonal anti-5-methylcytosine antibodies. Concentrations of IL-18 in cell culture supernatants were determined by commercial ELISA kit. Effects of maximum concentrations of genistein and SAMe on global DNA methylation were assessed using one-way ANOVA and post hoc LSD tests to determine the differences between groups. Effects of different concentrations of genistein and SAMe on IL18 production were evaluated using test for linear trend.
Results There was a significant difference in RASF global DNA methylation between control cultures and cultures treated with SAMe and genistein (pANOVA =0.028). Post hoc test revealed significantly higher levels of DNA methylation in RASF treated with 100 ug/ml SAMe than in those treated with 20 ug/ml genistein (p=0.01). Increasing concentrations of SAMe were associated with significantly up-regulated IL18 production by RASF (p for linear trend =0.044). There was no effect of genistein on IL18 synthesis.
Conclusions Both SAMe and genistein may have an effect on global RASF DNA methylation. SAMe up-regulates IL18 production by RASF which indicates that non-selective enhancement of DNA methylation may potentially lead to increased joint inflammation in RA.
Disclosure of Interest None declared
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